ISOLATION OF ARCANOBACTERIUM HAEMOLYTICUM FROM PATIENTS WITH PHARINGITIS

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ISOLATION OF ARCANOBACTERIUM HAEMOLYTICUM FROM

PATIENTS WITH PHARINGITIS

Sofia Constantiniu

1

, Micaela Scripcaru

1

, Angela Romaniuc

1

, Mariana Dumbrava

1

,

Anca Nistor

1

, Paraschiva Onu

2

1. Institute of Public Health Iasi, 2. Health Centre “Nicolina” Iasi

Abstract. Arcanobacterium haemolyticum has been described as a rare etiologic agent in acute pharingotonsilitis in pediatric and young population.Four strains of A. haemolyticum were isolated from throat swabs of 3584 patients (10-26 years ) with acute pharingitis. The samples, collected in January 1997-January 2001, were plated on Mueller-Hinton agar with 10 % sheep blood and incubated 24-48 hours at 37 °C.

In three cases A.haemolyticum was the only one etiologic bacterial agent and in the forth case association of A.haemolyticum with S.aureus was present.

The identification of A.haemolyticum strains was based on cells and colonies’ morphology, cultural and biochemical characters.

Key-words: Arcanobacterium haemolyticum, isolation, pharingitis.

Rezumat. A.haemolyticum a fost semnalat, cu frecvenţă redusă, ca agent etiologic al faringoamigdalitelor acute la copii şi tineri. Autorii descriu 4 tulpini de A.haemolyticum izolate din exudate faringiene de la bolnavi cu faringoamigdalită acută .3484 probe, recoltate in perioada ianuarie 1997-ianuarie 2001,au fost insămânţate pe agar Mueller- Hinton suplimentat cu 10 % sânge de berbec şi incubate la 37 °C timp de 24– 48 ore.

La 3 din cazuri A.haemolyticum a fost unicul agent izolat iar la 1 caz a fost prezent în asociaţie cu S.aureus.

Tulpinile de A.haemolyticum au fost confirmate pe baza morfologiei celulelor bacteriene şi a caracteristicilor culturale şi biochimice.

Cuvinte – cheie : Arcanobacterium haemolyticum, izolare, faringită.

A.haemoliticum (Corynebacterium

haemoliticum) is an aerobic gram –

positive rod that was first described by

MacLean et al., in 1946, isolated in

persons with pharingitis and skin

infections. Recently, many studies

point out that A.haemolyticum has

been isolated from patients with

pharingitis and various other infections in

Europe, USA and Asia (1,2 ).

The taxonomical position of

A.haemolyticum caused many confusions

because it is a close phenotypical similarity

to Actinomyces pyogenes (C. pyogenes ), an

animal bacterial pathogen (1). From

1982 this species was classified as A.

haemolyticum by Collins et al., in a

new genus Arcanobacterium composed

of this single species (1,3, 4).

In the infection with A.haemolyticum

clinical most cases involve pharingitis

and / or tonsilitis and approximately

50% are exudative. Throat infections

are often accompanied by cervical

lymphadenopathy (1,2,3,5). Simptoms

(2)

resemble those of beta – haemolytic

streptococci or viral infection.

An erythematous morbilliform or

scarlatinal rush of trunk, neck or

extremities were associated with the

presence of A. haemolyticum. In addition,

central nervous system infections, sepsis,

endocarditis, osteomyelitis, dermatologic

and other infections were described

(1,5).

The aim of this study were to determine

the presence of A.haemolyticum in acute

pharingitis in children and young

population and to characterize the

A.haemolyticum isolated strains.

MATERIAL AND METHODS

3584 throat culture samples were

collected during January 1997–

January 2001 from patients with acute

pharingitis. The patients of 5–26 years

of age were seen in districtual or

Children and Students Hospital of Iasi

city.

The samples were plated on Mueller –

Hinton agar added with 10% sheep

blood and incubated for 24–48 hours

at 37 C in aerobic conditions.

The A. haemolyticum strains were

identified by the following tests : the

small colonies (< 1 mm diameter) with

incomplete beta–haemolysis on sheep

blood agar, Gram stain, catalase,

nitrate reduction, urease, gelatin

hydrolysis, reverse CAMP, DNase

test, fermentation of glucose, lactose,

sucrose, maltose, mannitol and xylose

– table 1. The tests Voges–Proskauer

diffusion method. They were tested to

penicillin, erythromycin, tetracycline,

gentamycin, cephalotin, ceftazidime,

ceftriaxone, chloramphenicol, and

thrimethoprrim– sulphamethoxazole.

RESULTS AND DISSCUSION

A.haemolyticum was isolated in 4

(0.1%) cases of all investigated

patients (10–26 years). In 3 cases this

species was the only bacterial

pathogen and in one case was

associated with S.aureus .

Clinical manifestations of A.haemolyticum

pharyngitis were similarly with those of

streptococcal pharyngitis but the severity

of the disease varied.Sore throat and

pharyngeal erythema were always

present. Additional symptoms and signs

included fever, non–productive cough and

headache. Another symptoms and signs

such as skin rash, tonsillar exudates,

lymohadenopathy have also been

described. Erythema multiforme and

urticarial rash have also been found in

patients with A.haemolyticum pharyngitis

(1,6,7). In our investigation the frequency

of A. haemolyticum isolation was very

low. The isolation rate of this

pathogen from clinical throat

specimens varies from 0.07% to 1.3%

in non-age selected material (1). In

Romania were described 24 cases

confirmed with A.haemolyticum (6,7).

Epidemiological data suggest that

A.haemolyticum pharyngitis is

primarly a disease of adolescent and

young adults. Carlson P.; Coman et

(3)

on classical culture techniques.This

organism grows slow, with little (<1

mm diameter) and smooth/rough

colonies on blood agar and it is easly

overlooked in the bacteriological

laboratory. Around of the colonies

appears a narrow incomplete haemolysis

zone. Banck et al., considered that the

detection of beta- haemolysis

produced by this bacteria can be

facilitated by using special

double-layered human blood agar plates (3 ).

The identification of A.haemolyticum

was made by classical biochemical

tests presented in table 1 and 2. The

catalase and gelatin hydrolysis

differentiated this species of

Corynebacterium sp. and Actinomyces

pyogenes. A catalase negative,

gram-positive rods which is reverse CAMP

and DNase positive tests is very

important for the identification of

A.haemolyticum. The inclusion of

bacteria in genus and species have

been confirmed by the acid production

test from maltose, lactose, sucrose,

xylose and mannitol .

Many authors succesfully used the

API Staph and API Coryne systems

for testing carbohydrate fermentation

by A.haemolyticum (1,6,8).

This species can be divided in smooth

and rough biotypes by biochemical

tests. The majority smooth biotype

strains fermented sucrose and/or

trehalose in API Staph but not

produced beta-glucoronidase. The

rough biotype strains produced

beta-glucoronidase but not fermented

sucrose and trehalose.

All our A. haemolyticum strains were

susceptible to penicillin, erythromycin,

gentamycin,chloramphenicol, cephalotin,

cephtazidime,cephtriaxone.2 strains were

resistant to tetracycline and the

thrimethroprim-sulphamethoxazole had

no activity in vitro. In majority

studies, the A. haemolyticum strains

were susceptible to the antibacterial

agents recommended for treatment of

streptococcal tonsillitis, such as penicillin,

oral cephalosporins, erythromycin and

clindamycin (1,3,6, 8).

CONCLUSIONS

y A.haemolyticum was isolated from

0.1% of the throat specimens of

10-26 year old patients with acute

pharingitis, using sheep blood agar

on which this species grows as little

smooth or rough colonies.

y The identification of this organism

was possible using: Gram stain,

catalase, reverse CAMP, gelatin

hydrolysis and the capacity of acid

formation from glucose, maltose,

sucrose, xylose, mannitol .

y All A.haemolyticum strains were

susceptible to penicillin, cephalosporins,

macrolides, tetracycline and resistant to

thrimethoprim-sulphamethoxazole.

(4)

Table 1. Differential characteristics of A. haemolyticum and other bacterial groups Acid from Bacterial group Catal ase M ot ili ty Nitrat reducti on Ureas e Gel ati n hy dr olys is C arb ohyd ra te ferm ent ati on Gl ucose Esculin hydr olyz ed Mal tos e L acto se Su cr os e Xyl os e Mann itol Corynebacterium sp. + - V V V + + - V - V - - Arcanobacterium haemolyticum (C. haemolyticum) - - - - - + + - + + V - - Actinomyces pyogenes (C. pyogenes) - - - - + + + - V + V + V Listeria monocytogenes + + - - - + + + + V V - -

+, positive; -, negative; V, variable reaction.

Table 2. Principal characteristics of A. haemolyticum

Test Reaction Test Reaction β-haemolysis Catalase Nitrate reduction Urease Gelatin hydrolysis Esculin hydrolysis + - - - - - Reverso CAMP DNase Glucose Maltose Sucrose Xylose + + + + V -

(5)

REFERENCES

1. Carlson P: Arcanobacterium haemolyticum infections. Academic Dissertation, University of Helsinki, Finland, 1995, 1. 2. Mackenzie A., Fuite L.A., King J.,Allen

U.,MacDonald N.,Diaz-Mitoma F.: Incidence and pathogenicity of Arcanobacterium haemolyticum during a 2-year study in Ottawa. Clin. Infect. Dis., 1995,21, 1, 177.

3. Banck G., Nyman M.: Tonsilitis and rash associated with Corynebacterium haemolyticum. J.Infect. Dis.,1986,154, 1034.

4. Collins M.D.,Jones D.,Schofield G.M.: Reclassification of “Corynebacterium

haemolyticum” (McLean, Liebow &

Rosenberg) in the genus Arcanobacterium gen. nov. as Arcanobacterium haemolyticum nom.rev., comb.nov.

5. Coman G., Pânzaru C., Dahorea C.: Isolation of Arcanobacterium haemolyticum from the throat swab in children. Bacteriologia (Bucuresti ), 1996, 41, 3-4,145.

6. Koneman W.E., Allen D.S., Janda M.W., Schreckenberger C.P., Winn C.W.-The aerobic Gram –positive bacilli. In Diagnosis Microbiology, 4th edition, J. B. Lippincott Comp., Philadelphia, 1992, 498.

7 Dorobat O.,Erscoiu S., Burtea M.: Pharingitis due to Arcanobacterium haemolyticum. Bacteriologia (Bucuresti), 1996, 41, 3-4, 141.

8 Linder R.: Rhodococcus equi and Arcanobacterium haemolyticum: Two “ Coryneform” bacteria increasing recognized as agents of human infection. Emerg. Infect. Dis. 1997, 3, 2, 1.

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